Nucleic acid-based detection technologies play an important role in disease diagnosis, drug detection, environmental monitoring and criminal investigation, and there has been a continued need for new detection technologies which have excellent sensitivity and specificity and which are convenient to use. A nucleic acid-based detection technology, which has been typically used in the prior art, is based on a molecular beacon which is a U-shaped DNA probe labeled with a fluorophore and a quencher. This technology comprises confirming whether a fluorescent signal is produced by the conformational change of the molecular beacon due to the presence of a target nucleic acid (Tyagi et al., Nature biotechnology, 14:303-308, 1996; Masuko et al., Nucleic Acids Research, 26:5409-5416, 1998). Because this technology can rapidly analyze a target nucleic acid without isolating nucleic acids, it is widely used, and various types of molecular beacon-based nucleic acid analysis technologies have been developed (Song et al., Angewandte Chemie-International Edition, 48:8670-8674, 2009; Wu et al., Biosensors & Bioelectronics, 26:3870-3875, 2011; Yeh et al., Nano Letters, 10:3870-3875, 2011). However, the above-mentioned molecular beacon-based analysis technology has a shortcoming in that, because a target nucleic acid and a molecular beacon react at a ratio of 1:1 to generate a fluorescent signal, it is difficult to achieve high sensitivity.
In recent years, for the purposes of overcoming this problem and developing a detection sensor having excellent sensitivity, attempts have been made to use enzyme as a tool for signal amplification. A typical example includes a system in which an enzyme labeled with a detection probe for detecting a target nucleic acid and with an inhibitor is introduced (Gianneschi et al., Angewandte Chemie-International Edition, 46: 3955-3958, 2007; Saghatelian et al., Journal of the American Chemical Society, 125: 344-345, 2003; Pavlov et al., Journal of the American Chemical Society, 127: 6522-6523, 2005; Niemeyer et al., Angewandte Chemie-International Edition, 49: 1200-1216, 2010). In this system, the presence of the target nucleic acid blocks the inhibitory function of the inhibitor through hybridization with the detection probe, and the restored enzymatic activity generates a high fluorescent signal. However, this technology has shortcomings in that it requires a process of labeling the enzyme with the inhibitor, is time-consuming, and requires great skill. Furthermore, it has a shortcoming in that the labeling process itself may cause a conformational change of the enzyme to reduce the activity of the enzyme. In addition, this technology has a limitation in that it is hardly used as a versatile technology for detection of various target nucleic acids, because it is required to prepare an enzyme-inhibitor complex for each target nucleic acid. Thus, it is demanded to develop a label-free, enzyme-based universal for detection of various target nucleic acids.
Accordingly, the present inventors have made extensive efforts to overcome the above-described problems occurring in the prior art and to develop a highly sensitive enzyme-based detection and quantification system which does not need to be labeled with an inhibitor and which can be universally used for the detection and quantification of various target nucleic acids. As a result, the present inventors have found that the use of a DNA aptamer comprising a single-stranded DNA that specifically recognizes a target nucleic acid enables the following in a selective and accurate way, thereby completing the present invention: (1) detection and quantification of a target nucleic acid in switch-off mode; (2) detection and quantification of a target nucleic acid in switch-on mode; (3) detection and quantification of a target molecule in switch-on mode; (4) detection and quantification of target BER enzyme activity in switch-on mode; and (5) detection and quantification of a target nuclease in switch-on mode.